Bioequivalence and Bioavailability Forum

Bonjour chimiste!

❝ hello sir helmutz,

Not interested in opinions of other members of the forum?

❝ […] i would like to know how to calculate my LIQ frome un infernal Cmax, without taking into account the formula of 5%, is there any other mathematic relation between Cmax as pk parrameter and LIQ as analytical parrameter.

You should be able to quantify 5% (or lower) of C_max in order to demonstrate that there are no residual concentrations of previous administrations in periods >1. In a crossover study you have to avoid carry-over which requires a sufficiently long washout.
It is the job of the pharmacokineticist (not yours) to design the study. Hint: Never plan with the average half-life from the literature. Always assume longer ones (see this presentation, slides 65/66).
The bioanalytical method should be reliable and reproducible for the intended use. Sometimes people forget the last part and reach too far. It’s not necessary to come up with a “perfect” method – it must only be fit for purpose. Goalposts:

• ULOQ
  • Highest expected C_max in any subject.
    
  • Validate also dilution with blank matrix or analysis of an aliquot in case you find concentrations >ULOQ.
• LLOQ
  • AUC_0–t/AUC_0–∞ ≥80%.
    
  • ≤5% (individual!) C_max.
• (In)accuracy/(im)precision ≤20% at the LLOQ and ≤15% above.

❝ 2/ i would like to know how can i calculate the internal standard concentration used in validation from an ULOQ ?

Not sure what you mean. Can you reword/explain?

❝ 3/IAM working acctually on hplc and lc ms ms dosage methode of amoxicilline in human plasma , iam juste in begining and i need some of your precious recommandations if it is possible ?

Some hints from the method we used in my CRO (250 ng/mL – 31 µg/mL):

• Blood sampling / stability
  • Anticoagulant EDTA(K₃), Na-, or Li-heparin.
    
  • No stabilization necessary.
  • Centrifuge within 15 minutes after sampling or
    put in an ice bath and centrifuge within one hour.
• Long-term storage
  • At -20 ℃ stable for two months.
    
  • At -20 ℃ degradation ~25% after nine months.
    
  • At -70 ℃ stable for nine months.
• Stability of plasma at room temperature
  • Stable for four hours.
    
  • 6% degradation after 24 hours.
• Stable in three freeze/thaw-cycles.
  
• Stability of extracts (in our SFE-method 50% CH₃OH)
  • At room temperature for 48 hours.
    
  • At -20 ℃ for at least one week.
• Aqueous stock solution (1 mg/mL) at -70 ℃ stable for at least three months.

Good luck in developing/validating your method!

❝ N//B excuse me for my english iam doing my best because i usually communicate in frech.

No problem. Few native speakers here. translate is not that bad.